Ion gradients established by Na, K-ATPase in the nonpigmented ciliary epithelium (NPE) are vital to the secretion of aqueous humor. We seek to understand the mechanisms that control Na,K-ATPase activity. Regulation of Na,K-ATPase activity in the NPE is a point of control for the aqueous secretion mechanism. Our working hypothesis is that Na,K-ATPase activity, and thus AH secretion, is subject to regulation by Src family tyrosine kinases (SFK). NPE cells are rich in Na,K-ATPase as well as NO synthase and nitric oxide (NO) donors slow aqueous humor secretion. Studies this far suggest locally generated NO reduces NPE Na,K-ATPase activity. Pilot studies indicate NO activates one or more SFKs in the NPE. In AIM 1 we will test the hypothesis that SFK activation is a step in the pathway for NO regulation of Na,K-ATPase activity in the NPE. By regulating Na,K- ATPase activity and thus aqueous secretion, NO may serve an autocrine function. Preliminary studies show elevated IOP triggers NO generation. Carbonic anhydrase inhibitors (CAIs) reduce the rate of aqueous humor secretion. Although CAIs work by limiting the availability of cellular bicarbonate, their mechanism of action on aqueous secretion is obscure because they are just as effective in species that concentrate bicarbonate in the aqueous humor and those that do not. We have data that suggest bicarbonate transport inhibition activates a pathway that leads to SFK activation and inhibition of Na,K-ATPase activity. In AIM 2 we will test the hypothesis that interference with cellular bicarbonate regulates Na,K-ATPase activity by a SFK-dependent mechanism in the NPE. Drugs that slow aqueous formation are a proven way to lower IOP in individuals that have primary open angle glaucoma. The number of persons with glaucoma is increasing and in the future there will be a growing need for novel, affordable glaucoma drugs. The studies proposed here will improve our understanding of regulatory mechanisms for aqueous humor formation and may identify targets to consider in the search for new drugs to reduce aqueous production. PUBLIC HEALTH RELEVANCE: Ion gradients established by Na, K-ATPase in the nonpigmented ciliary epithelium (NPE) are vital to the secretion of aqueous humor. We seek to understand the mechanisms that control Na,K-ATPase activity because regulation of Na,K-ATPase activity in the NPE is a point of control for the aqueous secretion mechanism. Our working hypothesis is that Na,K-ATPase activity, and thus AH secretion, is subject to regulation by Src family tyrosine kinases (SFK). We will test whether SFK activation is a step in the pathway for nitric oxide regulation of Na,K-ATPase activity in the NPE. We also will test whether interference with cellular bicarbonate regulates Na,K-ATPase activity by a SFK-dependent mechanism in the NPE. Drugs that slow aqueous formation are a proven way to lower IOP in persons with glaucoma. The number of persons with glaucoma is increasing and in the future there will be a growing need for novel, affordable glaucoma drugs. The studies proposed here will improve our understanding of regulatory mechanisms for aqueous humor formation and may identify targets in the search for new drugs to reduce aqueous production.